Baseline Biopsy Specimens Research Articles

A pilot trial of personalized neoantigen pulsed autologous dendritic cell vaccine as adjuvant treatment in hepatocellular carcinoma.

e16264 Background: The primary treatment for hepatocellular carcinoma (HCC) is radical surgery. However, HCC has a high propensity to recur postoperatively and no standard adjuvant therapies have been approved. This Phase I trial aims to evaluate the safety and efficacy of an adjuvant personalized neoantigen pulsed autologous dendritic cell vaccine (Neo-DCVac-02) in patients with HCC. Methods: Eligible patients had histologically confirmed high-risk HCC after radical surgery. After obtaining the necessary baseline biopsy specimens and PBMCs of appropriate quality for DNA and RNA sequencing, neoantigen prediction, screening and synthesis were performed. Subsequently, leukapheresis was performed and DCs were isolated and cultured. DCs were then pulsed with neoantigen peptides to produce Neo-DCVac-02. On day 0, patients were treated with cyclophosphamide at a dose of 250 mg/m2. The prepared Neo-DCVac-02 vaccine was administered subcutaneously to the axillary and inguinal regions bilaterally on day 1, followed by daily administration of GM-CSF at a dose of 0.075 mg for 5 days (days 2-6). The primary objectives of this study were to assess feasibility and safety/tolerability. Secondary objectives included evaluation of immune responses (via vaccine response: IFNγ ELISpot), relapse-free survival (RFS) and overall survival (OS). Results: A total of 32 patients provided informed consent to initiate the process of personalized neoantigen discovery, of which 26 (81%) met the requirements for successful product selection for clinical manufacturing. Feasibility was demonstrated with 13 patients receiving Neo-DCVac-02. 13 patients did not receive the neoantigen vaccine (7 due to progressive disease, 6 due to withdrawal of informed consent because of COVID-19). The treatment was well tolerated with no grade 2+ treatment-related adverse events (TRAEs). The most common Grade 1 TRAEs were injection site reactions (75%), rash (16.7%), fatigue (8.3%), neutropenia (8.3%), and headache (8.3%). Five of 13 evaluable patients showed increased antigen-specific T-cell activity and more than 50% of the neoantigen peptides in Neo-DCVac-02, and sustained cellular responses were observed up to one year. Increases in T-cell activation/co-stimulation seen after treatment with Neo-DCVac-02, as demonstrated by flow cytometric analysis, suggest immune priming. At a median follow-up of 29.7 months, median RFS was not reached, with 1- and 2-year RFS of 84.6% and 60%, respectively, and all patients are alive. Patients with a strong immune response had a longer median relapse-free survival (not reached) compared to patients with a weak immune response (17.6 months, P = 0.003). Conclusions: These data demonstrate that Neo-DCVac-02 is safe, well tolerated and capable of inducing durable antigen-specific lymphocyte immune responses that may correlate with delayed HCC recurrence. Clinical trial information: NCT04147078 .

Significance of Crypt Atypia in Barrett’s Esophagus: A Clinical, Molecular, and Outcome Study

The aim of this study was to characterize baseline morphologic features of crypts in nondysplastic Barrett's esophagus and correlate them with DNA content abnormalities and risk of progression to high-grade dysplasia (HGD) or esophageal adenocarcinoma (EAC). The morphologic features of nondysplastic crypts in baseline biopsy specimens from 212 BE patients (2956 biopsy specimens) were graded histologically using a 4-point scale (crypt atypia levels, 0-3). DNA content abnormalities were detected using flow cytometry. In patients who had dysplasia in their baseline biopsy specimens, dysplasia was associated significantly with increasing grades of crypt atypia in the background nondysplastic Barrett's esophagus (P < .001). In a subset of patients without dysplasia at baseline (N= 149), a higher grade of crypt atypia was associated with longer Barrett's esophagus segment length (5.5 vs 3.3 cm; P= .0095), and a higher percentage of cells with 4N DNA content (3.67 ± 1.27 vs 2.93 ± 1.22; P= .018). Crypt atypia was associated with the development of any neoplasia (low-grade dysplasia and HGD/EAC). Although no significant association was noted between the grade of crypt atypia and increased 4N, aneuploidy, or progression to HGD/EAC, only patients with grade 2 or 3 crypt atypia showed increased 4N, aneuploidy, or progression to HGD/EAC. Patients with Barrett's esophagus likely develop dysplasia via a progressive increase in the level of crypt atypia before the onset of dysplasia, and these changes may reflect some alteration of DNA content.

Metabolite Alterations and Interactions with Microbiota in Helicobacter pylori-Associated Gastric Lesions.

Metabolites and their interactions with microbiota may be involved in Helicobacter pylori-associated gastric lesion development. This study aimed to explore metabolite alterations upon H. pylori eradication and possible roles of microbiota-metabolite interactions in progression of precancerous lesions. Targeted metabolomics assays and 16S rRNA gene sequencing were conducted to investigate metabolic and microbial alterations of paired gastric biopsy specimens in 58 subjects with successful and 57 subjects with failed anti-H. pylori treatment. Integrative analyses were performed by combining the metabolomics and microbiome profiles from the same intervention participants. A total of 81 metabolites were significantly altered after successful eradication compared to failed treatment, including acylcarnitines, ceramides, triacylglycerol, cholesterol esters, fatty acid, sphingolipids, glycerophospholipids, and glycosylceramides, with P values of <0.05 for all. The differential metabolites showed significant correlations with microbiota in baseline biopsy specimens, such as negative correlations between Helicobacter and glycerophospholipids, glycosylceramide, and triacylglycerol (P < 0.05 for all), which were altered by eradication. The characteristic negative correlations between glycosylceramides and Fusobacterium, Streptococcus, and Gemella in H. pylori-positive baseline biopsy specimens were further noticed in active gastritis and intestinal metaplasia (P < 0.05 for all). A panel including differential metabolites, genera, and their interactions may help to discriminate high-risk subjects who progressed from mild to advanced precancerous lesions in short-term and long-term follow-up periods with areas under the curve (AUC) of 0.914 and 0.801, respectively. Therefore, our findings provide new insights into the metabolites and microbiota interactions in H. pylori-associated gastric lesion progression. IMPORTANCE In this study, a panel was established including differential metabolites, genera, and their interactions, which may help to discriminate high-risk subjects for progression from mild lesions to advanced precancerous lesions in short-term and long-term follow-up.

A pilot trial of personalized neoantigen pulsed autologous dendritic cell vaccine as adjuvant treatment in hepatocellular carcinoma.

e14658 Background: The primary means of treating hepatocellular carcinoma (HCC) is radical surgery. However, HCC has a high propensity to recur post-operatively, and no standard adjuvant therapies have been approved. Tumor neoantigens, which are epitopes derived from tumor-specific mutations, may be utilized in personalized vaccines to activate T cell responses. This trial aims to assess the safety and efficacy of personalized neoantigen pulsed autologous dendritic cell vaccine (Neo-DCVac-02) in patients with HCC. Methods: Eligible patients had histologically confirmed high-risk HCC after radical surgery. Upon obtaining the baseline biopsy specimens and PBMCs of appropriate quality for DNA and RNA sequencing, neoantigen prediction, screening, and synthesis were performed. Subsequently, autologous dendritic cells (DCs) were generated and pulsed with neoantigen peptides to produce Neo-DCVac-02.On day 0, patients were treated with cyclophosphamide at a dose of 250 mg/m2. The prepared Neo-DCVac-02 vaccine was administered subcutaneously on day 1, followed by GM-CSF administered daily for 5 days. The primary objectives of this study were to evaluate the feasibility and safety/tolerability. Secondary objectives included assessment of immune responses (via vaccine response: IFNγ ELISpot), recurrence-free survival (RFS), and overall survival (OS). Results: A total of 32 patients provided informed consent to initiate the process of personalized neoantigen discovery, of which 26 (81%) met the requirements for successful product selection for clinical manufacturing. And feasibility was shown with 13 patients receiving Neo-DCVac-02. Thirteen patients did not receive neoantigen vaccines (7 for progressive disease,6 for withdrawal of informed consent because of COVID-19). Treatment was well-tolerated with no grade 2+ treatment-related adverse events (TRAEs). The common grade 1TRAEs were neutropenia (8.3%), injection site reactions (75%), rash (16.7%), fatigue (8.3%), and headache (8.3%). The median follow-up to date is 19.30 (14.4-31.9) months, with median RFS and OS not yet reached and a 1-year RFS of 83%. Five of 13 evaluable patients showed elevated antigen-specific T-cell activity by IFNγ ELISpot for more than 50% of neoantigen peptides in Neo-DCVac-02, and persistent cellular responses were noted out to 1 year. Increases in activation/co-stimulatory of T cells seen after treatment with Neo-DCVac-02 revealed by Flow-cytometric analyses, suggest immune priming. Conclusions: These data demonstrate that Neo-DCVac-02 is safe, well tolerated, and able to generate durable antigen-specific lymphocyte immune responses. Early signs of clinical efficacy were observed in HCC patients. Clinical trial information: NCT04147078 .

Resistance of ground glass hepatocytes to oral antivirals in chronic hepatitis B patients and implication for the development of hepatocellular carcinoma.

Ground glass hepatocytes (GGHs) have been shown to predict the development of hepatocellular carcinoma (HCC). Type I GGH and type II GGH harbor hepatitis B virus (HBV) pre-S1 and pre-S2 deletion mutants, respectively. Whether anti-HBV therapy can inhibit the expression of GGHs and potentially reduce HCC development is explored in this study. Two sets of liver specimens were included: the first contained 31 paired biopsy specimens obtained from chronic HBV patients receiving oral nucleos(t)ide analogue (NA) treatment; the second contained 186 resected liver tissues obtained from HBV-related HCC patients receiving surgery: 82 received NA before surgery and 104 did not. Compared with the baseline biopsy specimens, type I (P=0.527) and type II GGH (P=0.077) were not significantly decreased after 48 weeks of NA treatment in the first set of patients. In the second set, despite suppression of viral load (P<0.001) and periportal necrosis (P=0.006) in treated patients, GGH (P=0.594), cccDNA (P=0.172) and serum pre-S mutants (p=0.401) were not significantly suppressed. A significant decrease of type I (P=0.049) and type II GGH (P=0.029) could only be observed in patients after long duration of treatment (median duration: 4.3 years). In the treated patients, the persisted type II GGH remained an independent variable associated with decreased local recurrence-free survival of HCC (P=0.019) as in non-treated patients (P=0.001). In conclusion, the persistence of GGHs could explain the residual risk of HCC development under anti-HBV treatment. Therefore, intrahepatic GGHs and pre-S mutant are potential additional targets for HCC prevention in patients already receiving anti-HBV treatment.

Stereological Quantification of Immune-Competent Cells in Baseline Biopsy Specimens From Achilles Tendons

Background: Limited data exist on the presence and function of immune-competent cells in chronic tendinopathic tendons and their potential role in inflammation and tissue healing as well as in predicting long-term outcome. Purpose: To quantify subtypes of immune-competent cells in biopsy specimens from nonruptured chronic tendinopathic Achilles tendons and healthy control tendons. In addition, to examine whether findings in baseline cell biopsy specimens can predict the long-term presence of Achilles tendon symptoms. Study Design: Cross-sectional and case-control study; Level of evidence, 3. Methods: Fifty patients with nonruptured chronic Achilles tendinopathy and 15 healthy participants were included. At time of inclusion, an ultrasound examination was performed immediately before an ultrasound-guided Achilles tendon biopsy specimen was obtained. Tissue samples were evaluated immunohistochemically by quantifying the presence of macrophages (CD68-PGM1+, CD68-KP1+), hemosiderophages (Perls blue), T lymphocytes (CD2+, CD3+, CD4+, CD7+, CD8+), B lymphocytes (CD20+), natural killer cells (CD56+), mast cells (NaSDCl+), Schwann cells (S100+), and endothelial cells (CD34+) using a stereological technique. A follow-up examination was conducted more than 4 years (range, 4-9 years) after the biopsy procedure to evaluate the long-term presence of Achilles tendon symptoms. Results: Macrophages, T lymphocytes, mast cells, and natural killer cells were observed in the majority (range, 52%-96%) of biopsy specimens from nonruptured chronic tendinopathic Achilles tendons. CD68-KP1+ macrophages (0.29% vs 0; P = .005) and CD34+ endothelial cells (3% vs 0.97%; P = .04) were significantly more numerous in tendinopathic tendons compared with healthy tendons. The presence of iron+ hemosiderophages was more frequently observed in biopsy specimens obtained from the group who was asymptomatic at follow-up compared with the symptomatic group (42% vs 5%; P = .02). Conclusion: This study provides evidence for the presence of immune-competent cells in the majority of biopsy specimens from nonruptured chronic tendinopathic Achilles tendons. Macrophages and endothelial cells were significantly more numerous in tendinopathic tendons than in healthy tendons. The presence of iron+ hemosiderophages in baseline biopsy specimens was associated with a good prognosis. Clinical Relevance: New insight into the role of immune-competent cells in chronic Achilles tendinopathy.

Hepatocyte Cytokeratin 7 Expression in Chronic Allograft Rejection

We examined hepatocyte cytokeratin 7 (CK7) expression in chronic allograft rejection (CR), a ductopenic condition in which this has not been systematically evaluated, in 20 patients with the clinicopathologic diagnosis of CR and age-, sex-, and native-disease-matched control subjects. We also studied baseline biopsy specimens from both groups. Three pathologists independently reviewed H&E- and CK7-stained sections, counting interlobular bile ducts (BDs) and portal tracts (PTs), noting the morphologic pattern of injury and scoring hepatocyte CK7 expression (0, none; 1+, rare; 2+, multifocal, predominantly periportal; 3+, extension into the lobule; 4+, diffuse). Mean BD/PT ratios and CK7 scores were calculated. The mean BD/PT ratio (0.58) and CK7 score (1.01) for the "CR, diagnostic" group were significantly different from all other group means (P < .05); no other comparisons were significant (P > .05). A CK7 score of 1 or more was observed in 9 (56%) of 16 CR specimens and in 3 (7%) of 41 remaining specimens. Hepatocyte CK7 expression is frequently noted in CR, and it would appear to reflect ductopenia. CK7 staining may be a useful diagnostic adjunct in evaluation of transplant liver biopsy specimens.

C4d deposition on peritubular capillary (PTC) in the protocol biopsy of ABO‐incompatible kidney transplantation under the treatment with anti‐CD20 antibody and without splenectomy

Abstract: For the desensitization of A/B antigens, we had developed and reported a new potent immunosuppressive treatment, which is the pre‐prescription of anti‐CD20 monoclonal antibody with mycophenolate mofetil and low‐dose steroid. Using this kind of desensitization therapy, splenectomy is not required at the kidney transplantation. Complement C4d deposition on peritubular capillary (PTC) in graft biopsy has been reported as a relatively reliable marker for humoral rejection. However, the C4d deposition was often observed in the graft biopsy of ABO‐incompatible kidney transplantation even with no rejection findings. The aim of this study was to examine the effect of this treatment on C4d deposition on PTC. Baseline and protocol graft biopsies obtained from 12 recipients of ABO incompatible kidney transplants were evaluated by light and immunofluorescence microscopy. To elucidate the involvement of classical and/or lectin pathway of complement cascades in C4d deposition, we examined the deposition of the initial activating proteins on PTC, IgG and IgM in the classical pathway and mannose‐binding lectin (MBL), H‐ficolin, L‐ficolin, MBL‐associated serine protease (MASP)‐1 and MASP‐2 in the lectin pathway. Three out of nine available baseline biopsy specimens showed diffuse C4d and IgM deposition on PTC. In the protocol biopsy, nine of 12 specimens revealed diffuse C4d deposition on PTC. Five of them had positive deposition of IgM and H‐ficolin on PTC, whereas the other initial proteins were not detected in all specimens. Apart from one case, the histological findings of the protocol biopsies were normal or borderline changes. Our study suggested that although the new treatment with anti‐CD20 antibody treatment and without splenectomy was clinically effective, it did not perfectly inhibit C4d deposition on PTC. It also confirmed the dual activation of both classical and lectin pathways in the process of C4d deposition on PTC in ABO‐incompatible transplantation.

Downregulation of the Cyclin-Dependent Kinase Inhibitor p27 kip1 Might Correlate with Poor Disease-Free and Overall Survival in Inflammatory Breast Cancer

Purpose The objective of this study was to evaluate whether p27 kip1 downregulation is a prognostic factor in patients with inflammatory breast carcinoma (IBC). Patients and Methods Fifty-eight patients with IBC were treated between January 1994 and July 2002. Median age was 49 years. Thirty-eight patients had baseline biopsy specimens. Patients received preoperative chemotherapy with FAC (5-fluorouracil/doxorubicin/cyclophosphamide; 34%) or FAC followed by a taxane (66%). All patients underwent mastectomies. All patients received radiation therapy and hormonal treatment when indicated. Expression level of p27 kip1 was evaluated by indirect immunoperoxidase procedure. The p27 kip1 was considered downregulated if nuclear staining was present in > 50% of the neoplastic cells. Results Thirty-two patients (84%) had p27 kip1-downregulated tumors, and 6 patients (17%) had p27 kip1-normal tumors. Six patients (16%) exhibited a pathologic complete response. At a median follow-up of 43 months, 25 recurrences (66%) and 27 deaths (71%) occurred. Patients with p27 kip1-downregulated tumors had fewer pathologic complete responses (9% vs. 50%; P = 0.03) and had lower 4-year recurrence-free survival (23% vs. 83%; P = 0.03) and overall survival rates (36% vs. 83%; P = 0.01). Conclusion The p27 kip1 deregulation manifested by low protein cellular concentration might represent an adverse prognostic marker in IBC and could provide a valuable tool for selecting treatment for this aggressive disease.

Roles of Iron and HFE Mutations on Severity and Response to Therapy During Retreatment of Advanced Chronic Hepatitis C

Iron overload may cause or contribute to hepatic injury and fibrosis. Mutations in the HFE gene may influence development or progression of chronic liver disease by increasing iron stores or modulating immune responses. The aim of this work was to assess the influence of HFE mutations and serum and hepatic measures of iron status on baseline features and response to lead-in therapy in subjects with advanced chronic hepatitis C enrolled in the Hepatitis C Anti-viral Long-term Treatment to prevent Cirrhosis (HALT-C) Trial. Entry criteria included an Ishak fibrosis score >2 and lack of iron overload (Scheuer iron grade <3+) according to local study pathologists. All baseline biopsy specimens were rescored by consensus of study pathologists, and detailed assessment of stainable iron was performed. Hepatic iron concentrations were measured on portions of 144 liver biopsy specimens. A total of 1051 out of 1145 subjects agreed to HFE mutational testing (C282Y, H63D, S65C). Thirty-five percent carried at least one HFE gene mutation. There were no significant differences in the prevalence of HFE gene mutations among subjects with fibrosis (35.5%) versus cirrhosis (32.9%). Thirty-three percent of subjects had end-of-treatment and 16% sustained virologic responses. Presence of HFE mutations, in particular the H63D variation, was associated with increased end-of-treatment (40% vs 29%, P = .0078) and sustained virologic responses (20% with HFE mutation vs 14% sustained virologic response without HFE mutation; P = .009). Although HFE mutations (especially the most frequent H63D mutation) are associated with increased iron loading, they are also associated with increased sustained virologic responses in US patients with advanced chronic hepatitis C.

Assessment of Fibrosis Progression in Untreated Irish Women With Chronic Hepatitis C Contracted From Immunoglobulin Anti-D

In 1996 we initiated a retrospective-prospective study in 184 untreated women infected in 1977 with chronic hepatitis C virus (HCV). To provide insight into the natural history of HCV, we determined liver fibrosis outcomes and any predictors of such. Baseline 1994 biopsy specimens (size, >or=15 mm; portal areas, >or=5) and sequential biopsy specimens were assessed by Ishak score for grade change (increase or decrease of >or=2 points) and stage progression or regression (increase or reduction of >or=1 point), the latter correlated with digital quantification of fibrosis percentage. No baseline biopsy specimens had cirrhosis, therefore all could potentially progress. Grade and stage scores decreased or increased significantly in 28% and 18% and 24% and 27% of patients, respectively. There was a positive correlation between baseline and sequential grade/stage scores (r = .39, P < .001), and between semiquantitative Ishak scores and fibrosis percentage (Spearman rho = .85; P < .01). Baseline alanine transaminase values (mean, 49 U/L; range, 23-363 U/L) correlated positively with changes in grade (r = .41, P < .01) and stage (r = .39, P < .01), and regression analyses indicated that baseline alanine transaminase value was a good predictor of such changes. Confounding variables (alcohol, smoking, and herbal and paracetamol [acetaminophen] use) did not correlate with histologic outcomes. In a follow-up study, 49% of patients showed no change in fibrosis, 24% showed regression, and only 27% showed progression, including 4 patients (2.1%) who developed stage 6 cirrhosis. Unidirectional sequential grade/stage concordance attested to biopsy sample reliability. Given the current age of these women in their fifth decade, some still may have a risk for more advanced liver disease, but for most of these patients it appears unlikely.

Fully automated assessment of inflammatory cell counts and cytokine expression in bronchial tissue.

Automated image analysis of bronchial tissue offers the opportunity to quantify stained area and staining intensity in a standardized way to obtain robust estimates of inflammatory cell counts and cytokine expression from multiple large areas of histopathologic sections. We compared fully automated digital image analysis with interactive digital cell counting and semiquantitative scoring of cytokine expression in terms of repeatability and agreement in bronchial biopsies in 52 patients with mild to moderate atopic asthma. Immunohistochemistry with antibodies against CD3, interleukin (IL)-4, IL-5, and interferon-gamma protein was performed on frozen tissue sections, using 3-amino-9-ethylcarbazole as chromogen and hematoxylin as counterstaining. IL-4 and IL-5 messenger RNAs were localized by in situ hybridization without hematoxylin staining. Separation of 3-amino-9-ethylcarbazole and hematoxylin-stained pixels was achieved by linear combination of red- and blue-filtered gray-scale images. Using baseline biopsy specimens, fully automated CD3+ cell counts showed perfect repeatability (r = 1.0) and a strong linear relationship with the interactive procedure (r = 0.98). Automated densitometry showed perfect repeatability (1.0) and a moderate to strong relationship with semiquantitative scoring of protein and messenger RNA expression (r = 0.43-0.89). Relationships between automated and semiquantitative assessments of changes in cytokine expression during 2 years of follow-up were moderate to strong (r = 0.40-0.84). We conclude that fully automated cell counts and automated densitometric analyses in bronchial tissue of patients with asthma are unbiased and help to reduce variability in inflammatory outcomes.

Reversible pulmonary trunk banding. II. An experimental model for rapid pulmonary ventricular hypertrophy

Objective: An experimental model with a reversible pulmonary trunk banding device was developed with the aim of inducing rapid ventricular hypertrophy. The device consists of an insufflatable cuff connected to a self-sealing button. Methods: The right ventricles of 7 young goats (average weight, 8.7 kg) were submitted to systolic overload and evaluated according to the hemodynamic, echocardiographic, and morphologic aspects. Baseline biopsy specimens were taken from the myocardium for microscopic analysis. The device was implanted on the pulmonary trunk and inflated so that a 0.7 right ventricular/left ventricular pressure ratio was achieved. Echocardiographic and hemodynamic evaluations were performed every 24 hours. Systolic overload was maintained for 96 hours. The animals were then killed for morphologic study. Another 9 goats (average weight, 7.7 kg) were used for control right ventricular weight. Results: The systolic right ventricular/pulmonary trunk pressure gradient varied from 10.1 ± 4.3 mm Hg (baseline) to 60.0 ± 11.0 mm Hg (final). Consequently, the right ventricular/left ventricular pressure ratio increased from 0.29 ± 0.06 to 1.04 ± 0.14. The protocol group showed a 74% increase in right ventricular mass when compared with the control group. Serial 2-dimensional echocardiography showed a 66% increase in right ventricular wall thickness. There was a 24% increase in the mean myocyte perimeter, and the myocyte area increased 61%. Conclusions: The device is easily adjustable percutaneously, enabling right ventricular hypertrophy in 96 hours of gradual systolic overload. This study suggests that the adjustable pulmonary trunk banding might provide better results for the 2-stage Jatene operation and for the failed atrial switch operations to convert to the double-switch operation.J Thorac Cardiovasc Surg 2002;124:999-1006

Basophils, eosinophils, and mast cells in atopic and nonatopic asthma and in late-phase allergic reactions in the lung and skin

Background: Previous studies used indirect methods to identify basophils in the bronchi in asthma, and the numbers were not compared with eosinophils and mast cells. Furthermore, differences in basophil numbers between atopic and nonatopic asthma at baseline and between late-phase skin and asthmatic reactions have not been previously documented. Objective: The basophil granule–specific mAb BB1 was used to identify basophils in (1) bronchial biopsy specimens from atopic asthmatic subjects and nonatopic asthmatic subjects and control subjects, (2) biopsy specimens from atopic asthmatic subjects before and after inhalational allergen challenge, and (3) late-phase skin reactions. Basophil numbers were compared with EG2+ eosinophils and tryptase+ mast cells. Methods: Cells were enumerated in bronchial and skin biopsy specimens by means of immunohistochemistry with the alkaline phosphatase-antialkaline phosphatase method. Results: There were elevated numbers of basophils in baseline biopsy specimens in atopic asthmatic subjects compared with atopic control subjects or normal control subjects, although eosinophils and mast cells were 10-fold higher. There was an intermediate number of basophils in nonatopic asthmatic subjects. Basophils increased after allergen inhalation, but again basophils were less than 10% of eosinophils. In contrast, basophils in cutaneous late-phase reactions were approximately 40% of infiltrating eosinophils. The peak of basophil accumulation was at 24 hours, whereas maximal eosinophil infiltration occurred at 6 hours. One third of cutaneous basophils had morphologic appearances suggestive of degranulation. Conclusion: Numerous basophils infiltrated cutaneous late-phase reactions in atopic subjects. However, this cell was not prominent in bronchial biopsy specimens of asthmatic subjects, either at baseline or after allergen challenge. (J Allergy Clin Immunol 2000;105:99-107.)

Baseline glomerular size as a predictor of function in human renal transplantation.

Nonimmune mechanisms have been implicated in chronic renal allograft injury. In experimental studies, a strong correlation exists between glomerular size and the degree of glomerular sclerosis that develops after subtotal nephrectomy. Therefore, we assessed the impact of glomerular maximal planar area (MPA) in baseline biopsy specimens of human renal allografts on later graft function. The MPA was measured, by point counting and by computer planimetry, in postperfusion biopsy specimens from 96 allograft kidneys from nonhypertensive donors that had functioned for at least 2 years. Clinical data were analyzed throughout a follow-up period averaging 7.46+/-2.46 years. Both methods produced equivalent estimates of MPA. MPA proved to be a strong predictor of late renal allograft function, with a significant correlation (P = 0.02 to P < 0.01) between MPA at baseline and later serum creatinine level and creatinine clearance, beginning at 6 months after transplantation and persisting through follow-up. Creatinine level at discharge and occurrence of rejection were also independent predictors, whereas donor age, gender and race, cold ischemia time, cadaveric versus living donor, delay in initial function, and HLA mismatch did not predict clinical outcome. Larger glomeruli at baseline, measured by a simple point-counting technique, provide an early predictor of risk for late allograft dysfunction and may identify a subpopulation of patients in whom treatment to prevent/ameliorate glomerular enlargement and/or hypertension may be efficacious.

Heterogeneity of glomerular size in normal donor kidneys: Impact of race

Glomerular size has been the subject of many studies and, in a number of settings, has a direct association with the development of glomerular sclerosis. However, the normal distribution of glomerular size has not been thoroughly evaluated in the general population in the United States. To address this issue, we analyzed the baseline biopsy specimens of 103 human donor kidneys to determine the maximal planar area (MPA) of the glomerular tuft in a heterogeneous human population. The MPA of each glomerulus was determined by measurement of sections through the vascular pole and/or origin of the proximal tubule, and was determined on each section by two methods: point counting and computer planimetry. There was very high agreement between these two methods. Multivariate analysis was used to identify significant correlates with MPA. Overall, younger donors had smaller glomeruli (P < 0.0001). Black donors had a larger MPA (23.4+/-8.6 mm2 x 10(-3)) than white donors (17.9+/-6.7 mm2 x 10(-3); P < 0.001), independent of donor age. MPA was not significantly different between genders. This heterogeneity in glomerular size may confound clinical studies if not recognized and may help explain differences in glomerular structure and function in response to injurious processes. (Am J Kidney Dis 1998 Jul;32(1):43-6)

Density and localization of lymphocytes with natural-killer (NK) cell activity in periodontal biopsy specimens from patients with severe periodontitis.

Natural killer lymphocytes (NKL) in gingival biopsy specimens of 13 patients with advanced periodontitis and 6 normal control subjects have been evaluated by the monoclonal antibody NC 1, anti-B4 and a double immunoenzymatic labeling technique, combining the peroxidase-anti-peroxidase (PAP) method with the alkaline-phosphatase-anti-alkaline-phosphatase (APAAP) technique. Mononuclear cells with NK-activity (NKA) were mainly accumulated in the upper to middle third of the periodontal pocket. The concentration of NC-1-positive cells (CNC) of the baseline biopsy specimens was significantly increased at p = 0.05 compared to the CNC after hygiene training. The statistical difference rose to p = 0.01 following scaling, root planing and curettage therapy. The healthy gingiva of control subjects showed an average of 1-2 NKL, and no statistical difference could be found when compared to the CNC of the biopsy specimens of patients after periodontal therapy. These data support the hypothesis that antibody-dependent cytotoxic immunoreactive lymphocytes in chronic inflammatory periodontal disease can be reduced by scaling and root planing therapy.